The proposed investigation is designed to extend the use in NMR studies of proteins of a general system, developed in this laboratory, for the production of samples with specific patterns of labeling with stable isotopes. The labeled samples will be used to probe aspects of protein dynamics. The two, structurally related, thioredoxins form E. coli and from bacteriophage T4 have been chosen for study. There are high resolution x-ray structures of both proteins, extensive data on the solution properties of the E. coli protein, mutants of the E. coli protein known to affect folding kinetics, and differential stability of the oxidized and reduced forms. The sequential assignment of the T4 protein will be completed following the methodology already successfully used on the E. coli protein: Spectral editing using residue-specific enrichment with 2H, 13C and 15N labeled amino acids and 2D NOESY and COSY spectra on the 75% randomly deuterated proteins. The latter samples are central to the sequential assignment procedures and to the future dynamic studies on both proteins. They yield spectra of comparable sensitivity to the natural abundance protein with substantial enhancement of resolution due to reduced natural linewidths and reduced passive spin coupling. Reduced spin diffusion renders these random fractionally deuterated samples clearly superior to those at natural abundance for quantitative NOE distance analysis. Procedures for chiral beta deuteration will be developed. When combined with 15N enrichment, such samples should serve to extend both assignment and NOE structure determinations. Unidirectional rate constants will be determined for both the redox and denaturation transitions via chemical exchange and magnetization transfer experiments with samples possessing selected alpha and sidechain protonated positions in a perdeutero background. Such experiments will potentially both detect and provide the structural information on folding intermediates.